Electrochemical characteristics of a hyperthermophilic enzyme in microdroplets stirred and heated by surface acoustic waves

Micro total analysis system (μTAS) is expected to be applied in various fields. In particular, since electrochemical measurement is inexpensive and easy, electrochemical measurement can be integrated with a microchannel. However, electrochemical detection sensitivity in a microchannel is lowered because the diffusion of the detection target is limited. In an ordinary electrochemical detection system, using a stirrer in a beaker can overcome limited diffusion. We previously proposed a new detection system that combines a microliquid solution agitation technology using surface acoustic waves (SAWs) with the μTAS. The SAWs function as microstirrers, thus making electrochemical detection possible by overcoming limited diffusion of the sample. However, when the solution is stirred by the SAWs, the temperature of the solution increases to 70°C due to vibrational energy. This leads to enzyme inactivation and impaired electrochemical response. Therefore, in this study, we used a hyperthermophile-derived enzyme. Temperature and electrochemical characteristics of the detection system using SAWs and a multi-copper oxidase (MCO) derived from the hyperthermophilic archaea Pyrobaculum aerophilum were studied. Laccase, which is an MCO derived from the thermophilic fungus Trametes versicolor, was also studied. We also characterized the enzyme-electrochemical reaction using SAWs by comparing the magnitude of the reduction current obtained using the two enzymes with different heat resistances. We observed an increase in the electrochemical response with the SAWs, without impaired enzyme activity. Thus, the use of a thermostable enzyme is suitable for the development of a biosensor that uses SAWs for agitation.